Mismatch Repair Proteins Research Articles

NTRK gene alterations were enriched in hepatoid or enteroblastic differentiation type of gastric cancer

AimsCurrently, the clinicopathological characteristics of gastric cancer (GC) with oncogenic NTRK alterations are not well known. Although NTRK fusion has been identified as prevalent in DNA mismatch repair protein deficient...

Enhanced Commendable Sensitivity and Specificity for MSI in Colorectal Cancer by a New PCR-HRM Based 8-Loci MSI Assay.

This study evaluated the performance of the PCR-HRM assay by comparing it with immunohistochemistry (IHC) for mismatch repair (MMR) proteins and the PCR capillary electrophoresis (PCR-CE) methods. A total of 224 patients with colorectal cancer participated in the study, with nearly half having mismatch repair deficiency (dMMR) tissues and the remainder possessing pMMR tissues. There was a 97.77% concordance between the PCR-HRM assay and IHC, and a 97.56% concordance between PCR-HRM and the PCR-CE assay. In comparison with IHC for dMMR proteins, the PCR-HRM demonstrated a sensitivity of 96.36% and a specificity of 99.12%. When juxtaposed with the PCR-CE assay, its sensitivity was 98.96% and specificity stood at 96.33%. The mutations observed in the microsatellite loci were uniformly distributed across all eight loci. Discrepant outcomes were more frequent in instances of MLH1 and PMS2 deficiency. Furthermore, the germline mutation status of MLH1, MSH2, PMS2, and MSH6 in 62 patients was ascertained using next-generation sequencing. All patients displaying MMR gene pathogenic mutations (N = 14) were identified as MSI-H by PCR-HRM, whereas those with MSS tissues (N = 43) did not exhibit MMR gene pathogenic mutations. Thus, the PCR-HRM method proficiently pinpoints tumors with verified germline MMR mutations, indicative of Lynch syndrome. Conclusively, the PCR-HRM assay emerges as a swift and congruent diagnostic tool for microsatellite instability, boasting commendable sensitivity and specificity in colorectal cancer.

Comparing Clinicopathological and Immunohistochemical Features of Colorectal Carcinoma between Young and Old Age Groups.

The incidence of colorectal carcinoma (CRC) is increasing among individuals younger than 50, and some studies suggest the presence of differences in CRC among old and young individuals regarding clinical and histopathological features. The aim of this study was to compare clinicopathological features, mismatch repair protein status, and expression of certain immunohistochemical stains between young and old groups. The study included 180 cases and found significant histological and immunohistochemical differences between the two groups. CRC in the young tends to be more right-sided and has a higher percentage of dMMR proteins, but less expression of p53 mutations. These features are commoner in Lynch syndrome, and more investigations to study the relationship between young-onset CRC and hereditary syndromes are needed. Young-onset CRC also tends to show higher expression of tumor cell PD-L1, which is an expected finding, as dMMR cases are more likely to be immunogenic. Two other significant differences are the higher percentage of mucinous carcinoma and the higher tumor grade in young-onset CRC. These two features suggest a more advanced disease with possibly worse outcomes; however, there is no difference in disease stage between the two age groups.

Comparison of antigenicity between frozen section vs non-frozen section tissue blocks: An immunohistochemical study of antibodies commonly used in gynecologic pathology.

Frozen section (FS) is a technique widely used intraoperatively to render a preliminary histopathologic diagnosis, allowing for immediate decisions at the time of surgery. We aimed to investigate potential variations in tissue antigenicity induced by rapid freezing in a variety of gynecologic tumor samples. A total of 177 FS and 177 non-frozen section (NFS) tissue slides were tested using a panel of immunostains commonly used in gynecologic pathology, including hormone receptors (estrogen receptor, progesterone receptor), HER2, mismatch repair proteins (MSH6, PMS2), programmed cell death 1 ligand 1 (PD-L1), p53, napsin A, and ɑ-methylacyl coenzyme-A racemase. Immunohistochemistry results were categorized as positive or negative, and positive cases were subsequently scored based on the distribution and intensity of the staining. Certain immunostains, such as HER2, PD-L1, and p53, were scored according to the established guidelines. The overall concordance between FS and NFS blocks was 87%; among the 13% of discrepant cases, most (10.7%) were classified as minor, with only quantitative differences without foreseeable clinical significance. In 2.3% of cases, there were major qualitative changes with potential impact on disease management. We concluded that FS tissue blocks may, in most cases, safely be used for immunohistochemical studies because most discrepant cases showed only minor differences in staining, with no anticipated clinical significance. Nevertheless, for certain markers, including HER2, p53, and PMS2, a NFS block is preferred when that option is available.

DNA mismatch repair defect and intratumor heterogeneous deficiency differently impact immune responses in diffuse large B-cell lymphoma

ABSTRACT Deficient (d) DNA mismatch repair (MMR) is a biomarker predictive of better response to PD-1 blockade immunotherapy in solid tumors. dMMR can be caused by mutations in MMR genes or by protein inactivation, which can be detected by sequencing and immunohistochemistry, respectively. To investigate the role of dMMR in diffuse large B-cell lymphoma (DLBCL), MMR gene mutations and expression of MSH6, MSH2, MLH1, and PMS2 proteins were evaluated by targeted next-generation sequencing and immunohistochemistry in a large cohort of DLBCL patients treated with standard chemoimmunotherapy, and correlated with the tumor immune microenvironment characteristics quantified by fluorescent multiplex immunohistochemistry and gene-expression profiling. The results showed that genetic dMMR was infrequent in DLBCL and was significantly associated with increased cancer gene mutations and favorable immune microenvironment, but not prognostic impact. Phenotypic dMMR was also infrequent, and MMR proteins were commonly expressed in DLBCL. However, intratumor heterogeneity existed, and increased DLBCL cells with phenotypic dMMR correlated with significantly increased T cells and PD-1+ T cells, higher average nearest neighbor distance between T cells and PAX5+ cells, upregulated immune gene signatures, LE4 and LE7 ecotypes and their underlying Ecotyper-defined cell states, suggesting the possibility that increased T cells targeted only tumor cell subsets with dMMR. Only in patients with MYC¯ DLBCL, high MSH6/PMS2 expression showed significant adverse prognostic effects. This study shows the immunologic and prognostic effects of genetic/phenotypic dMMR in DLBCL, and raises a question on whether DLBCL-infiltrating PD-1+ T cells target only tumor subclones, relevant for the efficacy of PD-1 blockade immunotherapy in DLBCL.

Histopathologic, Molecular, and Clinical Profiling of Lymphoepithelioma-like Carcinoma of the Bladder

Lymphoepithelioma-like carcinoma of the bladder (LELC-B) is a rare histologic subtype characterized by strong immune cell (IC) infiltrates. A better prognosis and favorable response rates to immune checkpoint inhibitors have been described. We aimed to characterize the molecular profiles and IC infiltration of LELC-B for a better understanding of its therapeutic implications. We identified 11 muscle-invasive bladder cancer cases with pure and mixed LELC-B. Programmed cell death ligand-1 (PD-L1) expression and mismatch repair proteins were evaluated using immunohistochemistry. We calculated the tumor mutational burden and characterized mutational profiles using whole-exome DNA sequencing data. Transcriptomic signatures were detected using the NanoString nCounter PanCancer IO360 Panel. Multiplex immunofluorescence of tumor microenvironment (PD-L1, PanCK, α-SMA, vimentin, CD45, and Ki67) and T cells (CD4, CD3, PD-1, CD163, CD8, and FoxP3) was used to quantify cell populations. All LELC-B cases were highly positive for PD-L1 (median tumor proportion score/tumor cell, 70%; range, 20%-100%; median combined positive score, 100; range, 50-100) and mismatch repair proficient and negative for Epstein-Barr virus infection. IC infiltrates were characterized by a high CD8+ T-cell count and high PD-1/PD-L1 expression on immune and tumor cells. LELC-B showed upregulation of signaling pathways involved in IC response. Most common mutations were found in chromatin remodeling genes causing epigenetic dysregulation. All LELC-B cases showed high tumor mutational burden with a median of 39 mutations/Mb (IQR, 29-66 mutations/Mb). In conclusion, LELC-B is a highly immunogenic tumor, showing strong upregulation of PD-1/PD-L1 and making immune checkpoint inhibitors a promising treatment option.

Molecular classification of endometrial carcinoma on endometrial biopsy: an early prognostic value to guide personalized treatment

ObjectiveMolecular features are essential for estimating the risk of recurrence and impacting overall survival in patients with endometrial cancer. Additionally, the surgical procedure itself could be personalized based on the...

Effect of neoadjuvant chemoradiotherapy with or without PD-1 antibody sintilimab in pMMR locally advanced rectal cancer: A randomized clinical trial

Neoadjuvant chemoradiotherapy (NACRT) was the standard treatment for patients with locally advanced rectal cancer (LARC) with proficient mismatch repair (pMMR) proteins. In this randomized phase 2 trial (ClinicalTrial.gov: NCT04304209), 134 pMMR LARC patients were randomly (1:1) assigned to receive NACRT or NACRT and the programmed cell death protein 1 (PD-1) antibody sintilimab. As the primary endpoint, the total complete response (CR) rate is 26.9% (18/67, 95% confidence interval [CI] 16.0%-37.8%) and 44.8% (30/67, 95% CI 32.6%-57.0%) in the control and experimental arm, respectively, with significant difference (p= 0.031 for chi-squared test). Response ratio is 1.667 (95% CI 1.035-2.683). Immunohistochemistry shows PD-1 ligand 1 (PD-L1) combined positive score is associated with the synergistic effect. The safety profile is similar between the arms. Adding the PD-1 antibody sintilimab to NACRT significantly increases the CR rate in pMMR LARC, with a manageable safety profile. PD-L1 positivity may help identify patients who might benefit most from the combination therapy.

Universal Lynch Syndrome Screening in Colorectal Cancer: A 5-Year Experience of a Portuguese Pathology Department.

Lynch syndrome (LS) is a prevalent genetic condition associated with colorectal cancer (CRC). Accurate identification of LS patients is challenging, and a universal tumor screening approach has been recommended. We present the methodology and results of universal LS screening in our hospital's Pathology Department. This retrospective study analyzed CRC tumors from a 5-year period (2017-2021). Immunohistochemistry was used to assess MMR protein expression, followed by BRAF V600E analysis and MLH1 promoter methylation. Statistical analysis examined associations between clinicopathologic variables MMR status and LS-suspected tumors. The study analyzed 939 colorectal carcinomas, with 8.7% exhibiting mismatch repair (MMR) deficiency, significantly lower than previous research. After applying the algorithm, 24 LS-suspected cases were identified, accounting for 2.6% of tested patients and 29.3% of MMR-deficient tumors. Our study establishes the feasibility of universal testing for all new cases of CRC in detecting individuals at risk for LS, even in the absence of clinical information. To gain a comprehensive understanding of the MMR status in our population, further investigations are warranted.

Whole-Exome Sequencing of Vulvar Squamous Cell Carcinomas Reveals an Impaired Prognosis in Patients With TP53 Mutations and Concurrent CCND1 Gains

Very little information is available on the mutational landscape of vulvar squamous cell carcinoma (VSCC), a disease that mainly affects older women. Studies focusing on the mutational patterns of the currently recognized etiopathogenic types of this tumor (human papillomavirus [HPV]-associated [HPV-A], HPV-independent [HPV-I] with TP53 mutation [HPV-I/TP53mut], and HPV-I with wild-type TP53 [HPV-I/TP53wt]) are particularly rare, and there is almost no information on the prognostic implications of these abnormalities.Whole-exome DNA sequencing of 60 VSCC and matched normal tissues from each patient was performed. HPV detection, immunohistochemistry (IHC) for p16, p53, and mismatch repair proteins were also performed. Ten tumors (16.7%) were classified as HPV-A, 37 (61.7%) as HPV-I/TP53mut, and 13 (21.6%) as HPV-I/TP53wt. TP53 was the most frequently mutated gene (66.7%), followed by FAT1 (28.3%), CDKN2A (25.0%), RNF213 (23.3%), NFE2L2 (20%) and PIK3CA (20%). All the 60 tumors (100%) were DNA mismatch repair proficient. Seventeen tumors (28.3%) showed CCND1 gain. Bivariate analysis, adjusted for International Federation of Gynecology and Obstetrics stage, revealed that TP53 mutation, CCND1 gain, and the combination of the 2 alterations were strongly associated with impaired recurrence-free survival (hazard ratio, 4.4; P < .001) and disease-specific survival (hazard ratio, 6.1; P = .002). Similar results were obtained when p53 IHC status was used instead of TP53 status and when considering only HPV-I VSCC. However, in the latter category, p53 IHC maintained its prognostic impact only in combination with CCND1 gains. All tumors carried at least one potentially actionable genomic alteration. In conclusion, VSCCs with CCND1 gain represent a prognostically adverse category among HPV-I/TP53mut tumors. All patients with VSCCs are potential candidates for targeted therapy.

Optimizing Immunohistochemistry Reporting in Endometrial Cancer.

The management of endometrial cancer involves a multidisciplinary team (MDT) approach, with immunohistochemistry playing an important role in management and prognosis. Markers investigated include estrogen receptor (ER), progesterone receptor (PR), tumor protein 53 (p53), and mismatch repair(MMR)protein. Additionally, polymerase epsilon (POLE) mutations indicate treatment-responsive tumors, often with excellent prognosis. We aimed to improve the reporting of immunohistochemistry and introduce POLE testing, with a sustainable change in the long-term management of endometrial cancer at Royal Cornwall Hospitals Trust (RCHT). An initial sample of 53 patients with endometrial cancer from 2022 was analyzed. Endometrial biopsy reports were reviewed for immunohistochemistry reporting, with delays of reporting over 10 days documented. In initial results, a mean of 15.5% of cases failed to report p53 (12/53), ER (9/53), PR (10/53), and MMR (2/53). The interventions implemented in February 2023 were an immunohistochemistry proforma, the introduction of POLE testing, and departmental presentations. Data was re-collected between March and September 2023. After the project, there was a 100% (39/39) rate of reporting immunohistochemistry correctly. POLE testing was introduced to the department. In addition, the proforma developed is now standard practice in the reporting of endometrial cancer cases and is utilized in the gynae-oncology MDT meetings.

SWI/SNF Complex-Deficient Undifferentiated Carcinoma of the Pancreas: Clinicopathologic and Genomic Analysis

Inactivating alterations in the SWItch/Sucrose NonFermentable (SWI/SNF) Chromatin Remodeling Complex subunits have been described in multiple tumor types. Recent studies focused on SMARC subunits of this complex to understand their relationship with tumor characteristics and therapeutic opportunities. To date, pancreatic cancer with these alterations has not been well studied, although isolated cases of undifferentiated carcinomas have been reported. Herein, we screened 59 pancreatic undifferentiated carcinomas for alterations in SWI/SNF complex–related (SMARCB1 [BAF47/INI1], SMARCA4 [BRG1], SMARCA2 [BRM]) proteins and/or genes using immunohistochemistry and/or next-generation sequencing. Cases with alterations in SWI/SNF complex–related proteins/genes were compared with cases without alterations, as well as with 96 conventional pancreatic ductal adenocarcinomas (PDAC). In all tumor groups, mismatch repair and PD-L1 protein expression were also evaluated. Thirty of 59 (51%) undifferentiated carcinomas had a loss of SWI/SNF complex–related protein expression or gene alteration. Twenty-seven of 30 (90%) SWI-/SNF-deficient undifferentiated carcinomas had rhabdoid morphology (vs 9/29 [31%] SWI-/SNF-retained undifferentiated carcinomas; P < .001) and all expressed cytokeratin, at least focally. Immunohistochemically, SMARCB1 protein expression was absent in 16/30 (53%) cases, SMARCA2 in 4/30 (13%), and SMARCA4 in 4/30 (13%); both SMARCB1 and SMARCA2 protein expressions were absent in 1/30 (3%). Five of 8 (62.5%) SWI-/SNF-deficient undifferentiated carcinomas that displayed loss of SMARCB1 protein expression by immunohistochemistry were found to have corresponding SMARCB1 deletions by next-generation sequencing. Analysis of canonical driver mutations for PDAC in these cases showed KRAS (2/5) and TP53 (2/5) abnormalities. Median combined positive score for PD-L1 (E1L3N) was significantly higher in the undifferentiated carcinomas with/without SWI/SNF deficiency compared with the conventional PDACs (P < .001). SWI-/SNF-deficient undifferentiated carcinomas were larger (P < .001) and occurred in younger patients (P < .001). Patients with SWI-/SNF-deficient undifferentiated carcinoma had worse overall survival compared with patients with SWI-/SNF-retained undifferentiated carcinoma (P = .004) and PDAC (P < .001). Our findings demonstrate that SWI-/SNF-deficient pancreatic undifferentiated carcinomas are frequently characterized by rhabdoid morphology, exhibit highly aggressive behavior, and have a negative prognostic impact. The ones with SMARCB1 deletions appear to be frequently KRAS wild type. Innovative developmental therapeutic strategies targeting this genomic basis of the SWI/SNF complex and the therapeutic implications of EZH2 inhibition (NCT03213665), SMARCA2 degrader (NCT05639751), or immunotherapy are currently under investigation.

Effect of preoperative immune checkpoint inhibitors on reducing residual lymph node metastases in patients with gastric cancer: a retrospective study

Objective: To investigate the effect of immune checkpoint inhibitors on reducing residual lymph node metastasis in patients with gastric cancer. Methods: The cohort of this retrospective study comprised patients from Nanfang Hospital of Southern Medical University and the First Affiliated Hospital of Xiamen University who had undergone systemic treatment prior to gastrectomy with D2 lymphadenectomy and had achieved Grade 1 primary tumor regression (TRG1) from January 2014 to December 2023. After exclusion of patients who had undergone preoperative radiotherapy, data of 58 patients (Nanfang Hospital: 46; First Affiliated Hospital of Xiamen University: 12) were analyzed. These patients were allocated to preoperative chemotherapy (Chemotherapy group, N=36 cases) and preoperative immunotherapy plus chemotherapy groups (Immunotherapy group, N=22 cases). There were no significant differences between these groups in sex, age, body mass index, diabetes, tumor location, pathological type, Lauren classification, tumor differentiation, pretreatment depth of invasion by primary tumor, pretreatment lymph node stage, pretreatment clinical stage, mismatch repair protein status, number of preoperative treatment cycles, or duration of preoperative treatment (all P>0.05). The primary outcome measure was postoperative lymph node downstaging. Secondary outcomes included postoperative depth of invasion by tumor, number of lymph nodes examined, and factors affecting residual lymph node metastasis status. Results: Lymph node downstaging was achieved significantly more often in the Immunotherapy group than the Chemotherapy group (pN0: 90.9% [20/22] vs. 61.1% [22/36]; pN1: 4.5% [1/22] vs. 36.1% [13/36]; pN2: 4.5% [1/22) vs. 0; pN3: 0 vs. 2.8% [1/36], Z=-2.315, P=0.021). There were no significant difference between the two groups in number of lymph nodes examined (40.5±16.3 vs. 40.8±17.5, t=0.076, P=0.940) or postoperative depth of invasion by primary tumor (pT1a: 50.0% [11/22] vs. 30.6% [11/36]; pT1b: 13.6% [3/22] vs. 19.4% [7/36]; pT2: 13.6% [3/22] vs. 13.9% [5/36]; pT3: 13.6% [3/22] vs. 25.0% [9/36]; pT4a: 9.1% [2/22] vs. 11.1% [4/36], Z=-1.331, P=0.183). Univariate analysis revealed that both preoperative treatment regimens were associated with residual lymph node metastasis status in patients whose primary tumor regression was TRG1 (χ2=6.070, P=0.014). Multivariate analysis incorporated the following factors: pretreatment depth of invasion by primary tumor, pretreatment lymph node stage, pretreatment clinical stage, number of preoperative treatment cycles, and preoperative treatment duration. We found that a combination of immunotherapy and chemotherapy administered preoperatively was an independent protective factor for reducing residual lymph node metastases in study patients whose primary tumor regression was TRG1 (OR=0.147, 95%CI: 0.026-0.828, P=0.030). Conclusion: Compared with preoperative chemotherapy alone, a combination of preoperative immunotherapy and chemotherapy achieved greater reduction of residual lymph node metastases in the study patients who achieved TRG1 tumor regression in their primary lesions.

Histogenetic insights and genetic landscape of fibromatosis-like undifferentiated gastric carcinoma: a focused study

BackgroundThe aim of this study was to elucidate the histogenesis and genetic underpinnings of fibromatosis-like undifferentiated gastric carcinoma (FLUGC), a rare pathological entity.MethodThrough a detailed analysis of seven cases, including histopathological evaluation, CTNNB1 gene mutation screening, human epidermal growth factor receptor 2 (HER2) protein level quantification, and HER2 gene amplification assessment to identify the pathological and molecular characteristics of FLUGC.ResultsOf the seven patients in this study, five were male and two were female (age: 39–73 years). Four patients presented with lesions in the gastric antrum and three had lesions in the lateral curvature of the stomach. Histopathologically, over 90% of the tumor consisted of aggressive fibromatosis-like tissue, including proliferating spindle fibroblasts and myofibroblasts and varying amounts of collagenous fibrous tissues. Undifferentiated cancer cells, accounting for less than 10%, were dispersed among the aggressive fibromatosis-like tissues. These cells were characterized by their small size and were relatively sparse without glandular ducts or nested mass-like structures. Immunophenotyping results showed positive expression of CKpan, CDX2, villin, and p53 in undifferentiated cancer cells; positive expression of vimentin in aggressive fibromatosis-like tissue; positive cytoplasmic expression of β-catenin; and focal cytoplasmic positive expression of smooth muscle actin (SMA). Genetic analysis did not reveal any mutations in the CTNNB1 gene test, nor was there amplification in the HER2 gene fluorescence in situ hybridization (FISH) test. Additionally, the Epstein-Barr encoding region (EBER) of in situ hybridization was negative; and the mismatch repair (MMR) protein was positive. Programmed cell death-1 (PD-1) was < 1–5%; programmed cell death ligand 1 (PD-L1): TPS = 1–4%, CPS = 3–8.ConclusionThe study highlights the significance of CTNNB1, HER2, EBER, and MMR as pivotal genetic markers in FLUGC, underscoring their relevance for diagnosis and clinical management. The rarity and distinct pathological features of FLUGC emphasize the importance of accurate diagnosis to prevent underdiagnosis or misdiagnosis and to raise awareness within the medical community.

Investigate the relationship between Bacillus coagulans and its inhibition of chemotherapy-induced lung cancer resistance.

Lung cancer is a leading cause of death globally, with lung adenocarcinoma being the most common subtype. Despite advancements in targeted therapy, drug resistance remains a major challenge. This study investigated the impact of Bacillus coagulans on drug resistance in lung adenocarcinoma cells. The cells were pretreated with B. coagulans culture filtrate (BCCF), and functional assays were performed, including cell proliferation, cell cycle, apoptosis, and immunofluorescence staining. Results showed that BCCF induced cell cycle arrest at the S phase, reducing cell proliferation and suppressing drug resistance marker P-glycoprotein expression in BCCF-treated resistant cells rather than BCCF-treated control cells. Moreover, drug-resistant cells exhibited the ability for epithelial-mesenchymal transition, which could contribute to their necrosis through the iron-mediated cell death pathway upon BCCF treatment. Proteomic analysis identified downregulation of DNA mismatch repair protein PMS2 after BCCF treatment. These findings suggest that B. coagulans may modulate the DNA repair pathway, influencing drug resistance in lung adenocarcinoma cells. In conclusion, this study highlights the potential impact of B. coagulans on drug-resistant lung adenocarcinoma cells. Further investigation and understanding of the regulatory mechanisms by which B. coagulans modulates drug resistance in lung adenocarcinoma can aid in the development of more effective treatment strategies to improve the prognosis of lung cancer patients.

Modulation of miR-155-5p signalling via 5-ASA for the prevention of high microsatellite instability: an in vitro study using human epithelial cell lines

5-aminosalicylic acid (5-ASA) is a first-line treatment for maintaining colitis remission. It is a highly effective, safe, and well-tolerated drug with anti-inflammatory and chemo-preventive properties. While patients with primary sclerosing cholangitis (PSC) with concomitant ulcerative colitis are treated with 5-ASA, the molecular mechanisms underlying the drug’s chemo-preventive effects are not entirely understood. We previously reported that bile acids and lipopolysaccharide-induced miR-155 expression was associated with downregulating mismatch repair (MMR) proteins in CACO-2 cell lines. Therefore, in this investigation, a set of in vitro functional studies was performed to show the possible mechanisms behind the epigenetic relationship between miR-155 and 5-ASA’s prevention of high microsatellite instability (MSI-H). In transient transfection with miR-155Mimic, which behaves like endogenous miRNA, we confirmed the relationships between miR-155 and its target MMR in three human intestinal epithelial cell lines: CACO-2, NCM460D and HT-29. We have shown, for the first time, that 5-ASA modulates MLH1, MSH2, MSH6 in miR-155 transfected cells. These findings underline that chemoprotective 5-ASA therapy can effectively attenuate the expression of miR-155 and potentially prevent a development of MSI-H in a subset of colorectal cancers associated with PSC.

Systematic optimization of prime editing for the efficient functional correction of CFTR F508del in human airway epithelial cells.

Prime editing (PE) enables precise and versatile genome editing without requiring double-stranded DNA breaks. Here we describe the systematic optimization of PE systems to efficiently correct human cystic fibrosis (CF) transmembrane conductance regulator (CFTR) F508del, a three-nucleotide deletion that is the predominant cause of CF. By combining six efficiency optimizations for PE-engineered PE guide RNAs, the PEmax architecture, the transient expression of a dominant-negative mismatch repair protein, strategic silent edits, PE6 variants and proximal 'dead' single-guide RNAs-we increased correction efficiencies for CFTR F508del from less than 0.5% in HEK293T cells to 58% in immortalized bronchial epithelial cells (a 140-fold improvement) and to 25% in patient-derived airway epithelial cells. The optimizations also resulted in minimal off-target editing, in edit-to-indel ratios 3.5-fold greater than those achieved by nuclease-mediated homology-directed repair, and in the functional restoration of CFTR ion channels to over 50% of wild-type levels (similar to those achieved via combination treatment with elexacaftor, tezacaftor and ivacaftor) in primary airway cells. Our findings support the feasibility of a durable one-time treatment for CF.

Intriguing Insights From 100 Consecutive Colorectal Cancer Cases in Mid-Kerala: Sparse BRAF Gene Mutations and Mismatch Repair Deficiency (MMR-D).

Colorectal cancer (CRC) is among the most prevalent types of cancer globally. It is well established that the development of CRC primarily results from the sequential activation of oncogenes and the simultaneous inactivation of tumor suppressor genes. It has also been noted that after the initial oncogenic mutation, many subpopulations with different mutational profiles are created, causing heterogeneity among the tumors. This retrospective study analyzed 100 patients diagnosed with CRCthrough colectomy over an eighteen-month period at a tertiary referral center in mid-Kerala, India. Pathology records and histological slides were reviewed by two pathologists, and clinicopathological data were collected from pathology reports. Immunohistochemical analysis for BRAF mutation and possible microsatellite instability (MSI) (by mismatch repair (MMR) protein study) was conducted on tumor tissue blocks sent to an external center due to the lack of an automated platform at the hospital. The study utilized Roche's Benchmark XT platform for BRAF analysis and assessed MMR protein expression using antibodies for MLH1, MSH2, MSH6, and PMS2. The mean age of patients was 58.36 years, with a male predominance (58.0%). Most tumors were classified as T3 (71.0%, n-71) and T2/T4a (14.0% each, n-14), while nodal involvement included N0 (35.0%, n-35), N1 (26.0%,n-26), N2 (19.0%, n-19), and NX (20.0%, n-20). Histological examination revealed predominantly well-differentiated tumors (78.0%, n-78), with lymphatic invasion noted in 41.0% (n-41) and vascular invasion in 5.0% (n-5) of cases. Left-sided tumors predominated (33.0%, n-33), followed by rectal carcinoma (37.0%, n-37), and right-sided colon cancers (30.0%, n-30). Genetic profiling showed sparse BRAF mutations (1.0%, n-1) and MSI (1.0%, n-1), with some cases exhibiting loss of MMR proteins (MLH1, PMS2, MSH2, and MSH6) by immunohistochemistry (IHC). The study highlights the rarity of BRAF mutations in this cohort and emphasizes the diverse pathological and molecular characteristics observed. The discussion focuses on the implications of these findings, suggesting that CRC in this population exhibits unique clinicopathological features potentially influenced by factors beyond genetic mutations. Further multicentric studies are warranted to comprehensively explore these factors and refine risk stratification and treatment strategies for CRC patients in similar demographics.

Clinicopathological features and prognosis of sporadic mismatch repair deficient colorectal cancer

Objective: To investigate the clinicopathological characteristics and prognostic factors of sporadic mismatch repair deficient (dMMR) colorectal cancer. Methods: A total of 120 cases of sporadic dMMR colorectal cancer from July 2015 to April 2021 were retrospectively collected in Peking Union Medical College Hospital, Chinese Academy of Medical Sciences and Peking Union Medical College. Patients with Lynch syndrome; synchronous multiple colorectal cancers; preoperative anti-tumor treatments such as chemotherapy and radiotherapy; and those with incomplete follow-up information were excluded based on family history and next-generation sequencing (NGS) test results. Immunohistochemical stains were used to detect the expression of mismatch repair proteins, methylation-specific PCR for methylation testing, and fluorescent PCR for BRAF V600E gene mutation detection. The clinical and pathological data, and gene mutation status were analyzed. Follow-up was done to assess survival and prognosis including progression-free survival and overall survival rate. Results: Sporadic dMMR colorectal cancer occurred more frequently in the right side of the colon, in females, and in the elderly. Morphologically, it was mostly moderately-differentiated, and most patients had low-grade tumor budding. In terms of immunohistochemical expression, MLH1 and PMS2 loss were dominant, and there were age and location-specificities in protein expression. MLH1 methylation was commonly detected in elderly female patients and rare in young male patients; while MLH1 and PMS2 deficiency, and BRAF V600E mutation occurred more often on the right side (P<0.05). The 3-year and 5-year progression-free survival rates were 90.7% and 88.7% respectively, and the 3-year and 5-year overall survival rates were 92.8% and 90.7% respectively. Tumor budding status was an independent risk factor affecting patient recurrence (hazard ratio=3.375, 95% confidence interval: 1.060-10.741, P=0.039), patients with low-grade tumor budding had better prognosis, and those with medium or high-grade tumor budding had poor prognosis. Conclusion: For dMMR colorectal cancer patients, tumor budding status is an independent risk factor for recurrence.

Analysis of clinical characteristics of mismatch repair status in colorectal cancer: a multicenter retrospective study

BackgroundMicrosatellite instability (MSI) caused by DNA mismatch repair (MMR) deficiency is of great significance in the occurrence, diagnosis and treatment of colorectal cancer (CRC).AimThis study aimed to analyze the relationship between mismatch repair status and clinical characteristics of CRC.MethodsThe histopathological results and clinical characteristics of 2029 patients who suffered from CRC and underwent surgery at two centers from 2018 to 2020 were determined. After screening the importance of clinical characteristics through machine learning algorithms, the patients were divided into deficient mismatch repair (dMMR) and proficient mismatch repair (pMMR) groups based on the immunohistochemistry results and the clinical feature data between the two groups were observed by statistical methods.ResultsThe dMMR and pMMR groups had significant differences in histologic type, TNM stage, maximum tumor diameter, lymph node metastasis, differentiation grade, gross appearance, and vascular invasion. There were significant differences between the MLH1 groups in age, histologic type, TNM stage, lymph node metastasis, tumor location, and depth of invasion. The MSH2 groups were significantly different in age. The MSH6 groups had significant differences in age, histologic type, and TNM stage. There were significant differences between the PMS2 groups in lymph node metastasis and tumor location. CRC was dominated by MLH1 and PMS2 combined expression loss (41.77%). There was a positive correlation between MLH1 and MSH2 and between MSH6 and PMS2 as well.ConclusionsThe proportion of mucinous adenocarcinoma, protruding type, and poor differentiation is relatively high in dMMR CRCs, but lymph node metastasis is rare. It is worth noting that the expression of MMR protein has different prognostic significance in different stages of CRC disease.